Molecular Biology Week 7. Flashcards
1
Q
Describe the central dogma of molecular biology?
A
DNA to mRNA to proteins.
Trc Trl
2
Q
Describe Trc; translation:
A
This makes messenger RNA of the information in gene that codes for protein.
3
Q
Describe Trl; transcription:
A
This is the conversion of RNA message into protein at the ribosome.
4
Q
Is there a difference in the central dogma of molecular biology between prokaryotes and eukaryotes?
A
There is additional steps in eukaryotes; RNA processing and RNA transport.
5
Q
Where does trc and trl happen in bacteria?
A
In the cytoplasm.
6
Q
Where does trc and trl take place in eukaryotes?
A
Trc and RNA processing occur in the nucleus, trl occurs in the cytosol.
7
Q
What is a gene?
A
a gene is composed of specific base sequences organised in a way that allows DNA to be transcribed.
8
Q
What are the three components of a gene?
A
promoter, terminator and other regulatory sequences.
9
Q
What is a DNA relative to polypeptides ?
A
DNA is an informational storage unit that must be accessed to make the polypeptides of an organism.
10
Q
What are genes relative to the proteins of the cell and mRNA?
A
Genes are structural and code for proteins of the cell and mRNA is made from these during trc.
11
Q
What do some genes have that allows them to transfer and ribosomal RNA?
A
Some genes have an RNA product.
12
Q
What attracts the RNA polymerase?
A
The promoter structure.
13
Q
What are the (2) core promoter elements?
A
-10 (Pribnow box) and -35 bases before trc start site.
14
Q
What can make the promoting effect of the sequences stronger?
A
The more closely the promoter resemble the -10 pribnow box and -35 bases before the trc start site.
15
Q
What does RNA polymerase catalyse?
A
Trc.
16
Q
What are the multi-subunit (5) of the enzyme RNA polymerase?
A
Alpha 2, beta, beta prime, omega and sigma. ABBOT
17
Q
What does subunit (1) alpha 2 of the core enzyme RNA polymerase do?
A
The two alpha subunits assemble the enzyme and bind regulatory factors.
18
Q
What does the subunit (2) beta of the core enzyme RNA polymerase do?
A
This has the polymerase activity (catalyses the synthesis of RNA
19
Q
What does the subunit (3) beta prime of the core enzyme RNA polymerase do?
A
Binds to DNA (nonspecifically)
20
Q
What does the subunit (4) omega core enzyme RNA polymerase do?
A
The omega subunit restores denatured RNA polymerase to its functional form in vitro. It has been observed to offer a protective function to the B’ subunit. Promotes enzyme assembly.
21
Q
What are the two roles of the additional factor involved in the RNA polymerase holoenzyme? The sigma subunit (5)
A
The role of the sigma enzyme is to
1. Reduce the affinity of enzyme for non-specific DNA. (increase specificity)
2. Increase affinity for promoter sites.
(directs enzyme to the appropriate trc start)
22
Q
What happens to the sigma (5) factor after the promoter has been bound?
A
The sigma factor dissociates from the core enzyme.
23
Q
Name the three complex multi-subunit eukaryotic RNA polymerase types? How are they characterized?
A
They are characterized by how they synthesise.
- RNA polymerase I
- RNA polymerase II
- RNA polymerase III
24
Q
What is the most studied type of RNA polymerase?
A
Type II, this synthesises the precursors of mRNAs. A range of transcription factors are required for its binding to promoters.
25
What does the Type I of RNA polymerase synthesise?
A pre rRNA 45S.
26
What does the third type of RNA polymerase synthesise?
tRNAs, 5S rRNA and other small RNAs found in the nucleus and the cytosol.
27
What is the difference in the process of transcription between DNA and RNA.
Transcription follows the same base pairing rules as DNA replication to make RNA transcripts from the genes of the cells. However, uracil replaces thymine in the transcript that is produced.
28
What are the three steps in transcription?
1. Initiation
2. Elongation
3. Termination
29
Describe the first step in transcription; initiation?
Prokaryotes; RNA polymerase binds non-specifically to DNA. It is the sigma (5) that allows the enzyme to scan for promoter sites on the DNA.
30
What is the initial binding between the polymerase and a promoter referred to as?
A closed promoter complex. The DNA is not unwound.
31
What happens after the closed-promoter complex is bound?
The polymerase then unwinds approx. 12 bases of DNA around the initiation site to from an open-promoter in which single stranded DNA is available as a template for transcription.
32
What is transcription initiated by?
The joining of two free NTPs, after the addition of about the first 10 nucleotides, sigma (5) is released from the polymerase which is then moved along the template.
33
Is transcription more complex in eukaryotes in comparison to prokaryotes?
Yes.
34
Is RNA polymerase directly involved the recognition of the core promoter sequences?
No it does not directly recognise.
35
What mediates the binding of RNA polymerase forming a transcription initiation complex?
Transcription factors.
36
Can RNAP proofread?
Yes
37
How does RNAP facilitate elongation of the RNA?
P unwinds the template DNA ahead of it and rewinds the DNA behind it, maintaining an unwound region of about 17 bp in the region of transcription.
38
During elongation, the template strand of DNA is copied to RNA using complementary base pairing and insertion of the appropriate ------?
Ribonucleotides.
39
Can multiple RNAP transcribe a single DNA molecule simultaneously?
Yes
40
At termination, which two processes in bacteria lead to the release of the mRNA transcript.
Rho-dependent (has to bind to RNA polymerase) and Rho-independent (depends on secondary structures formed)
41
At termination in eukaryotes, it occurs in conjunction with t polyadenylation transcripts, what does this mean?
Polyadenylation is the addition of a poly(A) tail to a messenger RNA.
42
What are the four characteristics of the template strand ?
- serves as DNA template for transcription.
- addition of nucleotides occurs in the 5' to 3' direction by RNA polymerase.
- directionality of the template strand is in the opp direction 3' to 5'
- referred to as antisense strand, non-coding strand neg strand, transcribed strand.
43
What direction does the coding DNA strand run in?
- The 5' to 3' direction.
44
Does the coding DNA strand contain the same nucleotide sequence as the mRNA strand?
Yes, it is also referred to as the sense strand, non-template strand or positive strand.
45
What is the three differences between transcription in prokaryotes and eukaryotes?
1. Site of trc in cells.
2. Accessibility to RNA polymerase
3. Post trc modification of mRNA.
46
What is the difference between prokaryotes and eukaryotes in regards to post trc modification of mRNA?
- mRNA is not modified in prokaryotic cells.
In the eukaryotic cells the mRNA is :
- RNA splicing
- 5' end capping.
- addition of a polyA tail.
47
Where does translation occur?
In the cytplasm.
48
Where does transcription occur?
In the nucleus.
49
What are the three steps involved in the RNA processing in eukaryotes?
RNA splicing
RNA Capping
PolyA tail
50
What does RNA splicing involve?
This happens in EUKARYOTIC cells, trc usually produces a longer mRNA called pre-mRNA which must be processed to produce the mature mRNA.
51
What is the story with introns and exons?
Introns are removed by RNA splicing as RNA matures, meaning that they are not expressed in the final messenger RNA (mRNA) product, while exons go on to be covalently bonded to one another in order to create mature mRNA
52
Are introns transcribed?
Yes, but they are not translated.
53
What is splicing?
This is the process that involves the removal of the introns and linkage of the exons.
54
What is the splicesome?
This is a complex of snRNPs (small nuclear RNA and protein) that precisely remove introns from pre-RNA.
55
What is the particular intron RNA sequence within and at the intron-exon boyndary?
namely 5' splice site, branch site and 3' splice site.
56
What type of attachment does RNA capping involve? What does it involve and when does it occur?
It involves the covalent attachment of a modified guanosine to the 5' end of the mRNA. Occurs while transcription occurs after the first 20-25 nt have been transcribed.
57
What are the 2 functions of RNA capping?
- To allow for proper exiting of mRNA from the nucleus to the cytoplasm.
- CAP structure enables ribosome binding to mRNA.
58
Where is the PolyA tail located and what is it made up of?
At the 3' end of the most mature mRNA is a polyA tail of 100-200 adenine residues.
59
What are the one known and one unknown function of the polyA tails?
- PolyA tails leads to mRNA stability in the cytosol.
| - May also occur in bacteria, reason unknown.
60
What are the four main elements required for translation?
The genetic code, mRNA, tRNA and the ribosome.
61
What is a genetic code?
This is the relationship between the sequence of nts in the mRNA and the amino acids in the protein.
62
How is the genetic code read and in what numbers?
It is read in groups of three nts known as codons. The code constists of 64 different codons including 3 stop codons. It is almost universal and is degenerate.
63
What does translation involve?
This is the process to turning mRNA into protein.
64
Describe the bacterial mRNA untranslated regions:
It has a 5' untranslated region and a 3' untranslated region.The ribosome binding site is the 5' UTR.
65
Where is the start and stop codon of mRNA for bacteria?
The start codon or methionine is a few nts from RBS (ribosome binding site). The stop codon is at the end of the coding sequence followed by the 3' untranslated region.
66
What is the coding sequence?
This is a series of codons from the start to the stop codon that determines the sequence of amino acids in a polypeptide.
67
Describe the common structural features of tRNA in prokaryotic and eukaryotic tRNA.
- 3 step loops and a fourth stem with a single stranded region.
- anticodon at the end of stem-loop 2. A 3 base sequence complementary to a codon in the mRNA.
- 3' single stranded region is the acceptor sites for amino acid when tRNA is charged.
68
What is the function of the anticodon?
function of anticodons is to bring together the correct amino acids to create a protein, based on the instructions carried in mRNA. Each tRNA carries one amino acid, and has one anticodon.
69
How are different tRNA molecules names?
Named according to amino acid carried, there is more than one codon for an amino acid there is more than one corresponding tRNA.
70
How are tRNAs charged?
Enzymes called aminoacyl-tRNA synthetases catalyse the attachment of aa to tRNA
71
Give an example of the twenty different enzymes which charge the different amino acids?
ex: Alanyl-tRNA synthetase.
72
Describe the process of aminoacyl tRNA synthetase charging a tRNA:
A (1)specific amino acid and (2)ATP bind to aminoacyl-tRNA synthetase.
The amino acid is activated by the covalent binding of AMP, the pyrophosphate is released.
The correct tRNA binds to the synthetase.
The amino acid is covalently attached to the tRNA AMP is released.The charged tRNA is released.
73
Bacteria have one type of ribosome which?
Translates all of the mRNA in cytoplasm. Ribosomal proteins and rRNA are made and assembled in the cytoplasm.
74
Describe the ribosome of the eukaryotic cells?
These cells have distinct ribosomes found in various cell compartments.
75
Where are ribosomal proteins made in eukaryotic cells?
They are made in the cytoplasm and are moved into the nucleus to assemble with rRNA to make subunits. Assembly is then in the cytoplasm after the export of the subunits out.
76
What are the sedimentation coefficients of the bacterial and the eukaryotic cell in terms of small unit, large subunit and assembled ribosome?
30S 50S 70S
| 40S 60S 80S
77
During bacterial translation where does the mRNA lie between?
Within a space between the 30S and the 50S subunit.
78
Where does the synthesised polypeptide exit ?
Through a site in the 50S subunit of the bacterial translation process.
79
What are the three sites of the model of the ribosome structure?
Peptidyl, Aminoacyl and the exit site (PAE)
| In order: EPA
80
What are the three stages of translation?
Initiation, elongation and termination
81
What is the complex composed of at the start of the initiation of the process of translation?
A complex is formed between the (1) mRNA, the (2) first charged tRNA and the (3) ribosomal subunits.
This involved initiation factors and a supply of energy from the GTP (Guanosine triphosphate)
82
Why does initiation of translation in bacteria begin with the mRNA binding of the small ribosomal subunit?
This occurs due to a short ribosomal binding sequence near the 5' end of the mRNA pairing with a short sequence in the 16S rRNa.
83
Where is the start codon located at the start of translation, in the initiation phase?
It is usually located just a few nts downstream of this ribosomal binding site and the start codon of the mRNA is recognised and bound by an initiator tRNA.
84
Where does the large ribosomal subunit associate with the small subunit and the initiatior of the tRNA?
In the P site.
85
What are the three steps to the initiation of translation?
mRNA binds to the small ribosomal subunit, initiator tRNA binds to the start codon in mRNA and then the large ribosomal subunit (50S) binds. Initiation factors promote this binding.
86
In what three ways does initiation differ in eukaryotes from bacteria.
- instead of a RBS, eukaryotes have a 5' guanosine cap that is recognised by cap binding proteins to PROMOTE binding of mRNA to the small ribosomal subunit.
- start codon is at a VARIABLE distance from the RBS.
- Starting tRNA is METHIONINE as opposed to formyl methionine in bacteria.
87
What is the starting tRNA in eukaryotes and bacteria.
Eukaryote: Methionine
Prokaryote: Formyl-methionine.
88
What does elongation involve?
It involves covalent bonding of amino acid one at a time to creat polypeptide.
89
What are the three steps which follow during the addition of a new amino acid?
- polypeptide that has been synthesised is occupying the P site attached to a tRNA (polypeptide-tRNA)
- a charged tRNA binds to the mRNA in the A site of the ribosome-energy as GTP and ELONGATION factors are required for this binding.
- a polypeptide bond is formed between the aa in the A site and the growing p chain. The P chain is removed from the tRNA in the P site and transferred to the aa in the A site.
90
What is the peptidyl transfer reaction catalysed by?
A region of a 50S subunit known as peptidyltransferase centre.
91
Describe the process of elongation in translation of DNA synthesis? EPA
Translocation of the ribosome towards the 3' end of the mRNA by exactly one codon.
The tRNA in the P and A sites shift to E and P sites respectively.
The next codon in the mRNA is exposed in the unoccupied A site.
The uncharged tRNA exits the E site.
The next charged tRNA enters the empty A site and the same series of elongation steps take place.
Elongation continues until a stop codon moves into the A site.
92
What is a peptidyl transfer reaction?
This is a bond which forms between the polypeptide chain and the amino acid in the A site. The polypeptide is transferred to the A site.
93
What are the three stop codons which are not recognised by a tRNA?
UAA, UAG and UGA.
94
How are stop codons recognised?
They are recognised by release factors.
95
Describe the last steps in the termination of translation?
The completed polypeptide is in the P site and is released from the tRNA. The mRNA, ribosome subunits and the release factors dissociate.
96
Which direction does RNA polymerase synthesise each RNA transcript?
In the 5' to 3' direction, moving along the DNA template in the 3' to 5' direction.
97
How does the template strand differ during translation in relation to gene A B and C?
Gene A and B use the bottom strand and gene C uses the top strand
